Cell membrane receptors utilize numerous intracellular second messengers to propagate their signal into and through the cell. Individual receptors can activate many intracellular signaling pathways. An instrument capable of examining in real time either one specific signal pathway in detail or multiple pathways at the same time in a high throughput manner can be a valued asset to basic research. The Fluorometric Imaging Plate Reader (FLIPR 1) was originally designed to utilize fluorescent dyes to monitor changes in cell membrane potential. Subsequently the FLIPR1 was used to monitor rapid intracellular changes in calcium. The FLIPR1 has proven to be a valuable investigative tool for the investigators at the Medical University of South Carolina. Its age along with the lack of current support from the manufacturer has put the FLIPR1 in a condition bordering on obsolescence. The newest version of the FLIPR, FLIPRtetra, has expanded on the basic concepts of the original instrument. FLIPRtetra, features 96, 384 and 1536 well configurations, multi-wavelength kinetic readings, LED modules and user configurable excitation and emission filters, an optional adapter for addition of cell suspensions and enhanced luminescence sensitivity with the optional aequorin luminescence camera. With the user configurable LED excitation modules, the FLIPRtetra has expanded dye capabilities to include: ratiometric voltage sensing, mitochondrial membrane de-polarization, intracellular glutathione levels, reactive oxygen species and others. The FLIPRtetra will provide NIH-funded investigators a new tool to examine a myriad of intracellular signal systems and provide them with new information that will allow them to expand their research and knowledge.